Base pair sequence order in paired reads

[seqmonkey]

I have paired sequence reads which CASAVA splits into R1 and R2 (read 1 and read 2), and I would like to know how the base pairs are ordered in R2.

i.e.

Assuming that R1 is read 3'-5' and complementary to the template strand, is R2 then 5'-3' and still complementary to the template?

Or does Illumina software automatically convert it to 3'-5'?



Thanks.

[simonandrews]

The second read will be on the opposite strand to the first. Think of it like using the two PCR primers to sequence a PCR product since that's what you're effectively doing.

All mapping programs will expect to see reads this way so you don't normally need to do any manipulation before using the data for further processing.

[kmcarr]

I have paired sequence reads which CASAVA splits into R1 and R2 (read 1 and read 2), and I would like to know how the base pairs are ordered in R2.

i.e.

Assuming that R1 is read 3'-5' and complementary to the template strand, is R2 then 5'-3' and still complementary to the template?

Or does Illumina software automatically convert it to 3'-5'?



Thanks.

SeqMonkey,

DNA sequences, including reads from the Illumina, are always reported in the 5'->3' direction. There may be special circumstances in which they are reported in the 3'->5' direction but in those cases it will be made explicitly clear, usually by labeling the ends as written.